A cell goes to great lengths to ensure the proper biogenesis of proteins. During translation, the ribosome associates with multiple factors such as modification enzymes, chaperones and targeting complexes to guarantee that the final product is active, properly folded and correctly localized to the appropriate cellular location. Without these factors, the cell will be filled inactive, misfolded, aggregated and/or incorrectly allocated proteins. Such a scenario can result in the cell's malfunction and death, leading to many devastating diseases. Many studies have been carried out to understand the interaction of the ribosome with these molecular machineries;however, none of these studies look at how the ribosome handles and communicates with multiple complexes. In this proposal, we propose to study the interaction of a translating ribosome with two ribosome-associated factors that ensure proper biogenesis: the signal recognition particle (SRP) and the trigger factor (TF). SRP is a universally conserved targeting machinery and is responsible for delivering membrane and secretory proteins to the plasma membrane or to the endoplasmic reticulum. On the other hand, TF is a bacterial chaperone that aids in the folding of cytosolic and periplasmic proteins. SRP and TF have been shown to simultaneously bind to the ribosome, however, the molecular basis of how newly emerging proteins is funneled to either the SRP pathway or the TF pathway remains unclear. I propose to (1) test whether and how SRP and TF compete for binding to ribosome nascent chain complexes (RNCs);(2) determine if the recruitment of the SRP receptor helps exclude TF;and (3) test whether further elongation of the nascent chain favors TF interaction and removes SRP from the ribosome. Overall, the results of these proposed experiments will provide a comprehensive and quantitative molecular mechanism of how 'decision-making'is achieved by the ribosome and will serve as an important framework to understand the interplay of other ribosome- associated molecular machineries. PUBLIC HEALTH RELEVANCE: Upon translation of the protein, the ribosome interacts with multiple cellular factors to ensure that its final product is active, properly folded and correctly localized in the cell. In this proposal, we propose to study the interplay of two ribosome-associated complexes: the signal recognition particle and trigger factor. Results from the proposed experiments will help further our understanding of how the ribosome handles these molecular machineries and determine important concepts of how protein biogenesis is achieved.